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GSH Level of apoptotic vs non-apoptotic Jurkat Cells
Glutathione Detection Kit
Description:
Glutathione Detection Kit
Trade Name:
Chemicon (Millipore)
Product Overview:
Reduced glutathione (GSH) plays a role in the rescue of cells from apoptosis, by buffering an endogenously induced oxidative stress. Maintaining intracellular glutathione levels can prevent apoptosis. Glutathione levels diminish with age. Intracellular levels of glutathione (reduced GSH) are directly influenced by the presence of extracellular glutathione (oxidized GSSG). Apoptotic cells extrude glutathione in the reduced form at an early stage of the apoptotic process; for example, Jurkat, U937 and Hep G2 cells are known to exhibit this change (Ghibelli et al. 1998). Evidence has accumulated that extrusion of reduced glutathione alters the intracellular redox state, and this change may trigger the loss of mitochondrial membrane potential (Hall 1999). However, some cells, like HeLa and 3T3, apparently do not undergo cytosolic GSH change during apoptosis. The glutathione pathway is also involved in drug resistance of certain cancer types such as leukemia (Sargent et al. 1999).
Test Principle:
After inducing apoptosis, incubate cell lysates with monochlorobimane (MCB), a dye that has a high affinity for glutathione (MCB binds other thiols in the cell with lower affinity). The unbound dye is almost nonfluorescent, whereas the thiol-bound dye fluoresces blue (excitation = 380 nm; emission = 461 nm). Fluorescence can easily be detected using a fluorometer or a 96-well fluorometric plate reader.
Application:
The CHEMICON Glutathione Detection Kit is a quick method for detection of intracellular levels of reduced glutathione (GSH). Reduction in levels of GSH may be an early indicator of apoptosis.
Test Principle:
After inducing apoptosis, incubate cell lysates with monochlorobimane (MCB), a dye that has a high affinity for glutathione (MCB binds other thiols in the cell with lower affinity). The unbound dye is almost nonfluorescent, whereas the thiol-bound dye fluoresces blue (excitation = 380 nm; emission = 461 nm). Fluorescence can easily be detected using a fluorometer or a 96-well fluorometric plate reader.
Application:
The CHEMICON Glutathione Detection Kit is a quick method for detection of intracellular levels of reduced glutathione (GSH). Reduction in levels of GSH may be an early indicator of apoptosis.
Key Applications:
Activity Assay
Species Reactivity:
Mammals
Usage Statement:
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Kit or Assay Type:
- Apoptosis Assays
- Apoptosis Assays
Components:
- MCB Fluorochrome (Part No. 90127): 5 vials, 50 μg each.
- 4X Cell Lysis Buffer (Part No. 90130): One 16 mL bottle.
- 10X Wash Buffer (Part No. 90126): One 100 mL bottle.
- GSH Positive Control (Part No. 90146): 1 vial, 100 mg.
Storage Conditions:
Store kit materials at 4°C up to their expiration date.
Detection Methods:
Fluorescent
Materials Required but Not Delivered:
· Microcentrifuge
· Acetonitrile or DMF
· Adjustable volume pipettor with disposable tips
· Beakers, flasks, cylinders necessary for preparation of reagents
· Fluorescence plate reader with 380nm/460nm filter set
· Acetonitrile or DMF
· Adjustable volume pipettor with disposable tips
· Beakers, flasks, cylinders necessary for preparation of reagents
· Fluorescence plate reader with 380nm/460nm filter set